Hepatitis C virus (HCV) infection is a significant human health issue, accounting for 50% of liver transplants in the US, with approximately 170 million persons infected worldwide. The current standard of care therapy of PEGylated-interferon and ribavirin is effective in only about 50% of patients, with severe side effects and contraindications limiting patient compliance. New therapeutics targeting HCV protease have recently been approved, and additional therapies targeted the HCV NS5B RNA polymerase and the HCV NS5A accessory protein are in clinical trials. With each of these new therapies however, drug resistance is known to develop, and those new therapies have shown significant side effects. The experience with HIV therapy suggests that a truly effective regimen against HCV will consist of a cocktail of drugs against several HCV targets. HCV protein synthesis is initiated from the uncapped (+)strand RNA genome, utilizing a novel RNA structure in the 5'-untranslated region called an internal ribosome entry site (IRES). We have discovered two distinct chemical compound classes that have cross-genotype activity inhibiting HCV replication. The discovery process and preliminary mechanism of action studies indicate that the HCV IRES RNA is the likely target. Our Phase I STTR objectives are to further investigate structure-activity-relationships in each class through a combinatorial medicinal chemistry program. The compound scaffolds are readily accessible to a medicinal chemistry effort, allowing easy synthesis of many analogs. Certain hits with well-characterized activity can also be readily derivatized to determine the functional groups that are required for the low-micromolar potency against infectious HCV in cell culture. The Phase I objective is to comprehensively describe the structure- activity-relationships for both hit classes, and then synthesize and identify sub-micromolar leads that can lead to a Phase II program to develop novel clinical candidates for HCV therapy.